A NOVEL CDNA RESTORES REDUCED FOLATE CARRIER ACTIVITY AND METHOTREXATE SENSITIVITY TO TRANSPORT DEFICIENT CELLS

Mammalian cells accumulate reduced folates and methotrexate, a folate antagonist, through the reduced-folate carrier (RFC) (Goldman, I. D., Lichtenstein, N. S., and Oliverio, V. T. (1968) J. Biol. Chem. 243, 50 07-5017). This study describes the isolation and expression of a cDNA clone that restores RFC activity to human breast cancer cells defectiv e in this transporter. The cDNA codes for a peptide (mRFC1) of 58 kDa, whose hydropathy plot, resembling those of mammalian sugar transporte rs, predicts that it may be a member of a superfamily of transporter g enes. Transfection of methotrexate-resistant (MTX(R)) ZR-75-1 cells wi th an expression vector, pRFC1, that codes for this peptide restores t heir ability to accumulate methotrexate. Furthermore, transport of met hotrexate into pRFC1-transfected cells is blocked by a 10-fold molar e xcess of the reduced folate, 5-formyltetrahydrofolic acid, but is unaf fected by folic acid. The increase in methotrexate uptake that is obse rved in pRFC1-transfected MTX(R) ZR-75-1 cells reverses their resistan ce to this antitumor agent.